Large CAG/CTG repeat templates produced by PCR, usefulness for the DIRECT method of cloning genes with CAG/CTG repeat expansions.
We report here a simple method for generating large CAG/CTG repeat sequences. We have applied this method to clone the genomic sequence containing the CAG/CTG repeat and its upstream intronic sequence present in spinocerebellar ataxia type 3 or Machado-Joseph disease (SCA3/MJD) by a modified DIRECT method. With these modifications we have considerably simplified the generation of the repeat probe used to screen for anomalous bands. This method will facilitate the molecular approach to other genetic disorders where expansions of repeat sequences could be involved.
Pujana
M A
MA
Molecular Genetics Department, Medical and Molecular Genetics Center, IRO, Autovía de Castelldefels km 2.7, 08907, L'Hospitalet de Llobregat, Barcelona, Spain.
Volpini
V
V
Estivill
X
X
D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods*; D005260:Female; D006801:Humans; D017827:Machado-Joseph Disease / Q000235:genetics; D008297:Male; D018598:Minisatellite Repeats; D015345:Oligonucleotide Probes; D010375:Pedigree; D016133:Polymerase Chain Reaction / Q000379:methods*; D013132:Spinocerebellar Degenerations / Q000145:classification / Q000235:genetics; D018911:Trinucleotide Repeats
D015345:Oligonucleotide Probes